GPCRs constitute a large and diverse family of proteins that function to transduce extracellular stimuli into intracellular signals. They also represent nearly half of all pharmaceutical drug targets currently on the market today. Many GPCRs act as receptors for a variety of signaling molecules from small molecules to large proteins, including hormones, neurotransmitters, cytokines, and lipids. These proteins also act as receptors for environmental stimuli that are sensed as odors, tastes, pheromones, or light.
The GPCRs are integral membrane proteins that possess seven membrane-spanning helices with an extracellular N-terminus and an intracellular C-terminus. The GPCR is activated by the recognition of a ligand which leads to a conformational change in the receptor. The conformational change in the GPCR leads to the activation of a coupled G protein. Depending on the type of G protein to which the GPCR is coupled, a variety of downstream signaling pathways can be activated.
G proteins include heterotrimeric G proteins comprising three different subunits: the Gα subunit, the Gβ subunit, and the Gγ subunit. GPCRs transduce signals by catalyzing the dissociation of a coupled heterotrimeric G protein into a GTP-bound Gα subunit and a Gβ/Gγ subunit complex, each of which can independently initiate intracellular signaling events. There are many different Gα subunits, including Gαs, Gαt, Gαo, Gαi, Gαz, Gα15, Gαk, and Gαq. The different Gα subunits vary with regard to the GPCR that they may couple with and the signaling pathways they activate.
Several major signal transduction pathways are mediated by different Gα subunits, including stimulation (Gαs) or inhibition (Gαi) of the effector adenylyl cyclase, and activation of the effector phospholipase C (Gαq). Activation of the effector adenylyl cyclase by Gαs results in the modulation of the second messenger cyclic adenosine monophosphate (cAMP), which, in turn, can regulate the functions of protein kinase A and other proteins. The interaction of cAMP with protein kinases then leads to phosphorylation and activation of various transcription factors. Cell-based assays relying on transcriptionally controlled reporter genes could be well suited for high throughput screening and the identification of GPCR ligands. However, because each GPCR generally interacts with only one or a few types of Gα subunits, there is no universal assay that can be used for ligand or drug screening with all GPCRs. Furthermore, existing strategies for screening of GPCRs are sophisticated and expensive.